Transcranial magnetic stimulation enhances the specificity of multiple sclerosis diagnostic criteria: a critical narrative review

Background Multiple sclerosis (MS) is an immune-mediated neurodegenerative disease that involves attacks of inflammatory demyelination and axonal damage, with variable but continuous disability accumulation. Transcranial magnetic stimulation (TMS) is a noninvasive method to characterize conduction loss and axonal damage in the corticospinal tract. TMS as a technique provides indices of corticospinal tract function that may serve as putative MS biomarkers. To date, no reviews have directly addressed the diagnostic performance of TMS in MS. The authors aimed to conduct a critical narrative review on the diagnostic performance of TMS in MS. Methods The authors searched the Embase, PubMed, Scopus, and Web of Science databases for studies that reported the sensitivity and/or specificity of any reported TMS technique compared to established clinical MS diagnostic criteria. Studies were summarized and critically appraised for their quality and validity. Results Seventeen of 1,073 records were included for data extraction and critical appraisal. Markers of demyelination and axonal damage—most notably, central motor conduction time (CMCT)—were specific, but not sensitive, for MS. Thirteen (76%), two (12%), and two (12%) studies exhibited high, unclear, and low risk of bias, respectively. No study demonstrated validity for TMS techniques as diagnostic biomarkers in MS. Conclusions CMCT has the potential to: (1) enhance the specificity of clinical MS diagnostic criteria by “ruling in” true-positives, or (2) revise a diagnosis from relapsing to progressive forms of MS. However, there is presently insufficient high-quality evidence to recommend any TMS technique in the diagnostic algorithm for MS.


INTRODUCTION
Multiple sclerosis (MS) is an immune-mediated neurodegenerative and neuroinflammatory disease characterized by chronic central nervous system (CNS) degeneration with intermittent attacks of inflammatory demyelination and axonal damage (Reich, Lucchinetti & Calabresi, 2018).Mitigation of disease activity, disease progression, and disability accumulation requires early and correct diagnosis (McNicholas et al., 2018).To diagnose MS in a patient with a history suggestive of a demyelinating episode, clinicians must find evidence of lesion dissemination in space and time (Poser et al., 1983).In the 2017 McDonald criteria (Thompson et al., 2018), magnetic resonance imaging (MRI) and cerebrospinal fluid (CSF) oligoclonal bands aid the clinical history and exam in finding these features.The 2017 McDonald criteria are highly sensitive; however, their low specificity can lead to misdiagnosis, thus resulting in delayed diagnosis and unnecessary treatment in some individuals (Filippi et al., 2022;Gobbin et al., 2019;McNicholas et al., 2018).As such, it is desirable to discover biological markers (biomarkers) of disease activity in MS that have the sensitivity to identify subclinical lesions early in the disease course, while possessing high specificity for MS-related disease processes (Bielekova & Martin, 2004).
A biomarker is "an objectively measured indicator of normal biological processes, pathogenic processes, or … responses to a therapeutic intervention" (Bielekova & Martin, 2004).Biomarkers can aid diagnosis, classify the extent of disease, observe natural history, or monitor responses to treatments (Atkinson et al., 2001).The diagnostic utility of a biomarker is based on its performance against a reference standard (e.g., clinical-radiologic diagnostic criteria, histopathological diagnosis) (Adeniyi et al., 2016).A sensitive biomarker is one that yields a positive or abnormal result in a high proportion of individuals who have the disease (Adeniyi et al., 2016).A specific biomarker has a normal or negative result in a high proportion of individuals without the disease (Adeniyi et al., 2016).A diagnostic biomarker should ideally have both high sensitivity and high specificity (Adeniyi et al., 2016).Biomarkers are of interest in the broader biomedical literature because they can offer objective, biologically plausible information about a disease process that may go undetected by a patient (Strimbu & Tavel, 2010).In some cases, biomarker-based findings can precede clinical endpoints throughout the disease's natural history, leading to earlier diagnosis or signifying a change in the disease course (Andersen et al., 2021).In other cases, biomarker results can help distinguish a disease from other entities, leading to the correct diagnosis and targeted management (Hayes, 2015).In MS, a hypothetical diagnostic biomarker could be valuable to narrow the differential diagnosis in a patient with undifferentiated lesions on neuroimaging, or arrive at an earlier diagnosis in a patient with signs and symptoms suggestive of a demyelinating event (Bielekova & Martin, 2004;Paul, Comabella & Gandhi, 2019).
To aid diagnosis, a biomarker should balance the probability that a patient has MS and does not have an alternative diagnosis (Adeniyi et al., 2016;Atkinson et al., 2001;Richardson & Wilson, 2015).This problem is relevant in MS because the differential diagnosis is broad (Solomon, 2019;Solomon et al., 2023;Wildner, Stasiolek & Matysiak, 2020) and current diagnostic criteria-the 2017 McDonald criteria (Thompson et al., 2018) -are sensitive but not specific (Filippi et al., 2018(Filippi et al., , 2022;;Gobbin et al., 2019;van der Vuurst de Vries et al., 2018), resulting in a high rate of misdiagnosis (Dixon & Robertson, 2018;Solomon, 2019;Solomon, Naismith & Cross, 2019).A biomarker should likewise be reliable and valid; biologically plausible and clinically relevant; and practical and cost-effective (Adeniyi et al., 2016;Atkinson et al., 2001;Bielekova & Martin, 2004).A previous systematic review of TMS biomarker studies in MS, by this research group (Snow et al., 2019), highlighted cross-sectional relationships between various TMS techniques and MS clinical outcomes.However, the previous review did not directly address the role of TMS in MS diagnosis (Snow et al., 2019).Thus, the current critical narrative review aimed to explore the diagnostic accuracy of TMS techniques in MS.

SURVEY METHODOLOGY
This review followed the SANRA checklist (Baethge, Goldbeck-Wood & Mertens, 2019).The review is intended for clinicians and researchers with an interest in MS neurophysiology.

Search strategy
The search was planned by the entire study team and performed by a single author (NJS).
A single author (NJS) searched the PubMED, Embase, Web of Science, and Scopus electronic databases for studies published between January 1, 1985 (the first year of TMS publication) (Barker, Jalinous & Freeston, 1985) and February 28, 2022.The following search terms were adapted for each database: ("multiple sclerosis" (all fields) OR "clinically isolated syndrome" (all fields)) AND ("transcranial magnetic stimulation" (all fields)) AND (sensitiv Ã (all fields) OR specific Ã (all fields) OR "predictive value" (all fields) OR "likelihood ratio" (all fields) OR "odds ratio" (all fields) OR "risk ratio" (all fields) OR "hazard ratio" (all fields)).

Motor thresholds
Resting motor threshold (RMT) Lowest TMS stimulus intensity to elicit MEP with peak-to-peak amplitude of 50 mV in at least five of 10 consecutive trials, in resting target muscle (Rossini et al., 2015).Reported as % MSO.
Reflects the strength and size of the most excitable elements of the target muscle cortical representation, activity of glutamate its receptors (e.g., AMPA), and function of ion channels (e.g., VGSC) in cortical and spinal neuron populations (Rossini et al., 2015;Ziemann et al., 2015).

Motor evoked potential (MEP)
Deflection in EMG trace of target muscle following delivery of threshold or suprathreshold TMS pulse to target muscle cortical representation (Rossini et al., 2015;Ziemann et al., 2015).
Measured in active or resting muscle.MEP amplitude increases in sigmoidal relationship with TMS stimulus intensity.This stimulus-response curve requires incrementally increasing TMS stimulus intensity to examine corresponding increases in MEP amplitudes due to faster temporo-spatial summation at cortico-motoneuronal synapses (Rossini et al., 2015).Higher stimulus intensities improve synchronization of neuronal firing (Magistris et al., 1998).The stimulus-response curve indexes the excitability of the least to most excitable neuronal populations in the motor representation (Groppa et al., 2012;Ridding & Rothwell, 1997).Corticospinal conduction properties can be examined by observing MEP latency or waveform characteristics (Groppa et al., 2012;Rossini et al., 2015;Snow et al., 2019).
This method results in "re-synchronization" of corticospinal action potentials at the level of the peripheral motor neuron and overcomes trial-totrial variability in MEPs that is caused by phase cancellation and asynchronous firing of corticospinal motor neurons (Rossini et al., 2015).The main utility of TST is to examine corticospinal conduction deficits induced by demyelination (Chen et al., 2008;Vucic et al., 2023).Magistris et al. (1999) Silent periods

Corticospinal silent period (CSP)
Also known as contralateral silent period (CSP).
Quiescence in rectified EMG trace after MEP, when TMS is delivered during tonic contraction of target muscle (Rossini et al., 2015).CSP duration increases linearly with TMS stimulus intensity (stimulus-response curve) (Rossini et al., 2015).Reported as onset latency or duration of silent period.
The stimulus-response curve partly reflects gain and excitability characteristics of GABAergic inhibitory interneurons (Rossini et al., 2015;Ziemann et al., 2015).Short and long CSPs are mediated by GABA A -and GABA B -receptor activity, respectively (Rossini et al., 2015;Ziemann et al., 2015).The exact structural and functional mechanisms-including cortical versus spinal contributions-represent an area of intense scrutiny across the literature (Hupfeld et al., 2020;Škarabot et al., 2019;Yacyshyn et al., 2016).May index excitotoxicity (Snow et al., 2019).Tataroglu et al. (2003) Ipsilateral silent period (iSP) Suppression of background rectified EMG trace following a suprathreshold TMS pulse, during tonic contraction of the homologous muscle ipsilateral to the target motor area (Rossini et al., 2015).Reported as onset latency, duration, depth, or transcallosal conduction time.

Study screening
English, peer-reviewed journal articles of original studies were screened by a single author (NJS).Screening criteria were planned by the entire study team using the PICOS format (Schardt et al., 2007).
Intervention.Observational research that reported sensitivity and/or specificity.
Control.Healthy controls, free of neurologic or other disease; persons with alternative diagnoses.
Outcome.Sensitivity and/or specificity of any upper and/or lower extremity TMS technique.
Study.Cross-sectional or case-control studies comparing MS to control participants; or cohort studies following participants from symptom onset to diagnosis.

Data extraction
The approach to data extraction was planned by the entire study team and performed by a single author (NJS).The study team verified all transcribed data.
From study results, a single author (NJS) transcribed 2 × 2 contingency findings (Adeniyi et al., 2016;Glas et al., 2003;McInnes et al., 2018) (Table 2).Sensitivity was considered the percentage of participants with a diagnosis of MS, who exhibited abnormal TMS results (Sensitivity = True Positives ÷ (True Positives + False Negatives)).Specificity was considered the percentage of control participants (i.e., without a diagnosis of MS), who exhibited normal TMS results (Specificity = True Negatives ÷ (False Positives + True Negatives)).All studies provided sufficient information to determine sensitivity.In cases where insufficient information was available to determine specificity, only sensitivity was reported.Sensitivity or specificity estimates below 50% indicated that the TMS outcome performed worse than chance at ruling out or in MS, respectively.
When both sensitivity and specificity outcomes were available, the diagnostic odds ratio (DOR) was estimated (Glas et al., 2003).DOR was calculated as DOR = (True Positives ÷ False Negatives) ÷ (False Positives ÷ True Negatives) (Glas et al., 2003).Any DOR above 1.0 was associated with increased diagnostic accuracy (i.e., an increased odds that an abnormal TMS result was associated with diagnosis of MS) (Glas et al., 2003).DOR values were interpreted as trivial if < 1.68, small if 1.68-3.46,medium if 3.47-6.71,and large if > 6.71 (Chen, Cohen & Chen, 2010).When possible, 95% confidence intervals for sensitivity, specificity, and DOR were estimated using the methods outlined by Glas et al. (2003).

Critical appraisal
The approach to critical appraisal was planned by the entire study team and completed by a single author (NJS).The study team verified all critical appraisal findings.
Risk of bias.Risk of bias was assessed using the QUADAS-2 tool (Whiting et al., 2011), which evaluates studies' reporting in domains of participant selection, the index test (TMS), the reference standard (MS diagnostic criteria), and participant flow and timing.
Participant selection questions evaluated reporting of the participant selection process and the level of detail used to describe participant samples: i) Was a consecutive or random sample of patients enrolled?
ii) Was a case-control design avoided?
iii) Did the study avoid inappropriate exclusions?Index test questions assessed reporting of TMS data collection, analysis, interpretation, and summarization: i) Were the index test results interpreted without knowledge of the results of the reference standard (i.e., was blinding employed)?
ii) If a threshold was used, was it pre-specified?
Reference standard questions examined reporting of MS diagnostic criteria: i) Is the reference standard likely to correctly classify the target condition?
ii) Were the reference standard results interpreted without knowledge of the results of the index test (i.e., was blinding employed)?
Flow and timing questions appraised reporting of participant exclusions and the timing between MS diagnosis and TMS testing: i) Was there an appropriate interval between index test and reference standard?
ii) Did all patients receive a reference standard?
iii) Did all patients receive the same reference standard?iv) Were all patients included in the analysis?A single author (NJS) answered signaling questions as Yes/No/Unclear, to derive High/ Low/Unclear risk of bias for each domain.Based on the risk of bias from each domain, an overall risk of bias rating was assigned to each study (Sterne et al., 2019).
Biomarker validity.To explore whether studies provided sufficient evidence to justify TMS techniques as biomarkers for MS diagnosis, Bielekova & Martin's (2004) MS biomarker criteria were used.The Bielekova & Martin (2004) criteria classify biomarker studies according to MS-related pathophysiologic process, grade studies' methodologic quality, evaluate studies' clinical utility, and assess studies' clinical usefulness.
MS-specific pathophysiologic processes were classified as: i) Biomarkers reflecting alteration of the immune system, ii) Biomarkers of blood-brain barrier (BBB) disruption, iii) Biomarkers of demyelination, iv) Biomarkers of oxidative stress and excitotoxicity, v) Biomarkers of axonal/neuronal damage, vi) Biomarkers of gliosis, and/or vii) Biomarkers of remyelination and repair.
Methodologic quality of studies was based on the following questions: i) Are complete (raw) data provided?
ii) Was there an independent comparison to a reference standard or age-and sex-matched reference group?
iii) Was an appropriate spectrum of patients included (e.g., clinical subtypes, sample size)?iv) Were the methods used valid (e.g., data collection, processing, and analysis)?
v) Was there a processing and/or work-up bias (e.g., blinded processing and analysis)?
Clinical utility was evaluated according to the following criteria: i) Biological rationale (i.e., rational association with a pathogenic aspect of MS).
ii) Clinical relevance (i.e., positioned in the causal chain of pathological events leading to a meaningful clinical endpoint).
v) Correlation with disability/prognosis (i.e., relationship with disability accumulation over time).
Clinical usefulness was assessed against the following criteria: i) Sensitivity/specificity (i.e., sensitivity/specificity relative to reference standard).
ii) Reliability (i.e., consistency of a measurement across time or raters, probability of false-positive or false-negative results).
iii) Evaluation of a biomarker in epidemiological studies or natural history cohorts (i.e., establishing a statistical relationship between the biomarker and clinical endpoint in cross-sectional or longitudinal studies).
iv) Evaluation of a biomarker in proof-of-principle clinical trials (omitted).
A single author (NJS) applied ratings of Yes/No/Unclear to each criterion, to derive Yes/ No/Unclear ratings for each domain.Based on ratings in each domain, an overall validity rating (Yes/No/Unclear) was assigned to each study (Sterne et al., 2019).

Visual presentation of TMS outcomes
To visually compare the diagnostic accuracy of TMS techniques across studies, a single author (NJS) prepared a Forest plot of DOR point estimates and their 95% confidence intervals, organized by TMS outcome and study.DOR values were coded according to risk of bias rating.DOR values were interpreted as above (Chen, Cohen & Chen, 2010;Glas et al., 2003).

RESULTS
See Fig. 2 for study selection (Page et al., 2021).The authors identified 964 records after duplicate removal.The authors reviewed 187 full texts after title and abstract screening and retained 17 articles for data extraction and critical appraisal.

Participant characteristics
MS participant characteristics are detailed in Table 3, while control group characteristics are summarized in Table S2.Across all studies, there were 1408 MS participants (median n = 79, range = 44-162) and 690 control participants (median n = 34, range = 10-155).Few studies matched MS and comparison groups for age or sex.Most studies used the Poser criteria to diagnose MS (Poser et al., 1983), whereas no study employed the 2017 McDonald criteria (Thompson et al., 2018).Average disease or symptom duration ranged from 1-10 years (median 4.6 years).Median disability score, measured using the Expanded Disability Status Scale (EDSS) (Kurtzke, 1983) was 2.5 (range 1.5-6).

TMS findings
Table 4 and Fig. 3 summarize diagnostic accuracy findings of all TMS techniques studied.Table S3 highlights TMS methods and Table S4 provides an in-depth summary of TMS findings.Every study reported the sensitivity of the TMS techniques employed.Specificity and DOR could be gleaned from only eight studies (Beer, Rösler & Hess, 1995;Cruz-Martínez et al., 2000;Hess et al., 1987;Magistris et al., 1999;Mayr et al., 1991;Ravnborg et al., 1992;Schmierer et al., 2000;Tataroglu et al., 2003).Only central motor conduction time (CMCT) and MEP size (amplitude or area) were investigated by at least half of the studies reviewed.Therefore, only these techniques are discussed in detail below.

DISCUSSION
This review aimed to summarize the diagnostic accuracy and validity of TMS techniques to aid in the diagnosis of MS.Across all TMS techniques studied, there was modest sensitivity for MS at best.Of the few studies that evaluated specificity, only CMCT and MEP size (amplitude or area) were represented in enough studies to comment on overall diagnostic performance.Most studies had a high risk of bias and did not demonstrate validity for TMS biomarker use.

Diagnostic performance of TMS techniques
Recent reviews have discussed the role of TMS as a biomarker in MS (Alsharidah et al., 2022;Simpson & Macdonell, 2015;Snow et al., 2019;Ziemann et al., 2011).While there is optimism for using TMS to diagnose, monitor natural history, or assess treatment response in MS (Alsharidah et al., 2022), there is also hesitancy towards widespread clinical use of TMS due to lack of sufficient evidence and high risk of bias (Simpson & Macdonell, 2015;Snow et al., 2019).In the current review, studies failed to justify the validity of TMS techniques as biomarkers for MS diagnosis; however, some outcomes could help characterize corticospinal conduction loss throughout the disease course.TMS elicits corticospinal motor responses partly by stimulating axon terminals or axonal bends in superficial presynaptic layer II/III/V myelinated neurons at the gyral crown of the precentral gyrus (Siebner et al., 2022).Given MS is characterized by attacks of inflammatory demyelination and axonal damage (in the context of ongoing axonal degeneration) (Pachner, 2021;Reich, Lucchinetti & Calabresi, 2018), these processes can intuitively be characterized using TMS measures of CNS conduction (Vucic et al., 2023;Ziemann et al., 2011).For example, CMCT, MEPs, and triple stimulation technique (TST) could theoretically serve this role (Alsharidah et al., 2022;Simpson & Macdonell, 2015;Snow et al., 2019;Vucic et al., 2023;Ziemann et al., 2011).In the present review, only CMCT and MEP size had sufficient evidence to evaluate their diagnostic performance.
Only CMCT was sensitive for MS (median 75%), when combining findings from upper plus lower extremities.Both CMCT and MEP amplitude had high specificity for MS (median 100%).DOR was greatest and most consistent for CMCT of the lower extremities (median DOR 25.15).Both techniques were generally correlated with pyramidal function, cerebellar function, and EDSS; however, their associations with MRI findings (lesions, atrophy) were inconsistent.Both techniques had poor sensitivity for subclinical lesions, whereas CMCT was 96% specific for subclinical disease.CMCT was 75% sensitive for recovery from active disease but had negligible sensitivity for new disease activity.Lastly, CMCT was highly sensitive for SPMS and PPMS.McDonald criteria was as high as 100% (Gobbin et al., 2019).CMCT therefore has little additive value to enhance the sensitivity of MS diagnostic criteria.However, the greatest strength of CMCT would be its high specificity (median 100%), where the 2017 McDonald criteria was as low as 14% specific in one study (Gobbin et al., 2019).CMCT could reduce false-positive diagnoses (Schwenkenbecher et al., 2019), by "ruling-in" persons with corticospinal conduction loss who were identified as having MS as per the 2017 McDonald criteria.

Alternative utilities for TMS techniques
The current review also found a compelling role for TMS in characterizing MS natural history.For example, CMCT was highly sensitivity for both progressive MS and higher disability status.Given most of the evidence herein is cross-sectional, the authors cannot assign any causal relationship between corticospinal conduction deficits and disease progression or disability accumulation (Pachner, 2021).Nevertheless, CMCT could identify early neurodegeneration to help diagnose a transition to SPMS or revise the diagnosis of RMS to PPMS.The potential role of CMCT in identifying occult neurodegeneration would be especially important, given there are limited treatment options for progressive MS and the diagnosis of PPMS or conversion of RMS to SPMS requires evidence of chronic and irreversible disability accumulation (Hamdy et al., 2022;Thompson et al., 2018).To better establish whether CMCT could expedite the diagnosis of progressive MS subtypes will require more evidence from prospective longitudinal studies.

Limitations
Despite the novelty of this review in terms of addressing the diagnostic performance of TMS and critically appraising the TMS biomarker literature, several limitations should be noted.First, the 2017 McDonald criteria (Thompson et al., 2018) currently represents the gold standard of MS diagnosis.None of the articles reviewed used these criteria, and most used the Poser criteria (Poser et al., 1983).While the authors could not identify any past research that directly compared Poser and 2017 McDonald criteria, successive iterations of the McDonald criteria have generally been shown to diagnose MS earlier and more frequently than the Poser criteria (i.e., enhanced sensitivity) (Brownlee et al., 2015), but with compromised specificity and a higher rate of misdiagnosis (Tintoré et al., 2003).
Sensitivity and specificity of the Poser criteria have been estimated at 87% and 94%, respectively (Engell, 1988;Izquierdo et al., 1985), whereas sensitivity and specificity of the 2017 McDonald criteria range between 68-100% and 14-61%, respectively (Filippi et al., 2018(Filippi et al., , 2022;;Gobbin et al., 2019;van der Vuurst de Vries et al., 2018).In past work, the addition of CMCT did not enhance the sensitivity of the Poser criteria to increase MS diagnoses, but increased specificity and reduced MS misdiagnoses (Beer, Rösler & Hess, 1995).It presently is unclear how TMS techniques would perform in the context of the 2017 McDonald criteria, and this question should be addressed in future research.Next, this critical review is based on only 17 studies of a select few TMS techniques.Methods like paired-pulse and dual-coil TMS, or TMS-EEG, offer unique ways to explore intracortical excitability in excitatory, inhibitory, and neuromodulatory interneurons; other regions important to sensorimotor function (e.g., premotor cortex, supplementary motor area, cerebellum, somatosensory cortices); and distant non-motor regions (Rossini et al., 2015).Such techniques could characterize biologically plausible disease mechanisms not reviewed here, such as acute excitotoxicity or chronic neurodegeneration, linked to disease activity and progression, respectively (Chaves et al., 2019;Snow et al., 2019).
Lastly, given this is a critical narrative review, the results must be interpreted more carefully compared to a systematic review or meta-analysis.The studies included in the review are heterogeneous in terms of study design, sample size, participant characteristics, and TMS methods.The current approach to data synthesis and interpretation does not take study heterogeneity into account.Moreover, because the authors did not produce a single estimate for the diagnostic performance of each TMS technique, there is greater onus on the reader to interpret the findings.Nonetheless, the authors provide a protocol-driven review, following evidence-based methods for data extraction and critical appraisal.

CONCLUSIONS
MS is an immune-mediated neurodegenerative disease characterized by attacks of inflammatory demyelination and axonal damage, with variable but continuous accumulation of disability.Various TMS techniques can characterize conduction loss and axonal damage in the corticospinal tract.Most notably, CMCT could be a putative biomarker to: (1) enhance the specificity of the 2017 McDonald criteria by "ruling-in" true-positive MS diagnoses, (2) revise a diagnosis from RMS to PPMS, or (3) help arrive at an earlier diagnosis of SPMS.Herein, the authors summarized the current state of the literature and determined both a high risk of bias and poor justification for the validity of TMS techniques as diagnostic biomarkers in MS.In the future, more rigorous, prospective, longitudinal studies will be required, using comparisons to the 2017 McDonald criteria.
Craig S. Moore conceived and designed the experiments, analyzed the data, authored or reviewed drafts of the article, and approved the final draft.Michelle Ploughman conceived and designed the experiments, analyzed the data, authored or reviewed drafts of the article, supervision, Project Administration, and approved the final draft.

Figure 1
Figure 1 Simplified schematic of transcranial magnetic stimulation (TMS)-induced generation of motor evoked potential (MEP).(A) Pulse generator produces an electric current that is stored in, and rapidly discharged from, a large capacitor into the stimulating coil.(B) The insulated coil contains windings of copper, which generate a focal magnetic field from the electric current.(C) The magnetic field undergoes little attenuation from extracerebral tissues and painlessly induces an electric current in underlying layer II/III/V pyramidal neuron axons at the gyral crown of the primary motor cortex.(D) Activation of corticospinal pyramidal neurons elicits descending corticospinal volleys from the brain to the spinal cord by directly activating pyramidal tract neurons, or indirectly via interneurons that synapse onto the pyramidal tract/lateral corticospinal tract.(E) The descending corticospinal volley activates the target muscle, via stimulation of anterior horn cells, peripheral nerve, and motor unit.(F) The TMS-induces motor evoked potential (MEP) can be recorded via electromyography (EMG), with recording electrodes placed over the belly of the target muscle.(G) Analyzing the amplitude, latency, duration, and waveform characteristics provides information on the excitability and conduction characteristics of corticospinal pyramidal cells; the post-MEP corticospinal silent period (CSP) characterizes the excitability of corticospinal inhibitory interneurons.See refs:(Chaves et al., 2021;Rossini et al., 2015;Siebner et al., 2022;Snow et al., 2019;Spampinato et al., 2023) .Full-size  DOI: 10.7717/peerj.17155/fig-1

Table 5
Risk of bias assessment.